Localization of fibroblast growth factor-1 in cholinergic neurons innervating the rat larynx.

Cholinergic neurons in the dorsal motor nucleus of the vagus (DMNV) are particularly vulnerable to laryngeal nerve damage, possibly because they lack fibroblast growth factor-1 (FGF1). To test this hypothesis, we investigated the localization of FGF1 in cholinergic neurons innervating the rat larynx by immunohistochemistry using central-type antibodies to choline acetyltransferase (cChAT) and peripheral type (pChAT) antibodies, as well as tracer experiments. In the DMNV, only 9% of cChAT-positive neurons contained FGF1, and 71% of FGF1-positive neurons colocalized with cChAT. In the nucleus ambiguus, 100% of cChAT-positive neurons were FGF1 positive. In the intralaryngeal ganglia, all ganglionic neurons contained both pChAT and FGF1. In the nodose ganglia, 66% of pChAT-positive neurons were also positive for FGF1, and 90% of FGF1-positive ganglionic cells displayed pChAT immunoreactivity. Neuronal tracing using cholera toxin B subunit (CTb) demonstrated that cholinergic neurons sending their axons from the DMNV and nucleus ambiguus to the superior laryngeal nerve were FGF1 negative and FGF1 positive, respectively. In the nodose ganglia, some FGF1-positive cells were labeled with CTb. The results indicate that for innervation of the rat larynx, FGF1 is localized to motor neurons, postganglionic parasympathetic neurons, and sensory neurons, but expression is very low in preganglionic parasympathetic cholinergic neurons.     

Electrophysiological study on the effects of leptin in rat dorsal motor nucleus of the vagus

Immunoreactivity of leptin receptor (Ob-R) has been detected in rat dorsal motor nucleus of the vagus (DMNV). Here, we confirmed the presence of Ob-R immunoreactivity on retrograde-labeled parasympathetic preganglionic neurons in the DMNV of neonatal rats. The present study investigated the effects of leptin on DMNV neurons, including parasympathetic preganglionic neurons, by using whole cell patch-clamp recording technique in brain stem slices of neonatal rats. Leptin (30-300 nM) induced membrane depolarization and hyperpolarization, respectively, in 14 and 15 out of 80 DMNV neurons tested. Both leptin-induced inward and outward currents persisted in the presence of TTX, indicating that leptin affected DNMV neurons postsynaptically. The current-voltage (I-V) curve of leptin-induced inward currents is characterized by negative slope conductance and has an average reversal potential of -90 +/- 3 mV. The reversal potential of the leptin-induced inward current was shifted to a more positive potential level in a high-potassium medium. These results indicate that a decrease in potassium conductance is likely the main ionic mechanism underlying the leptin-induced depolarization. On the other hand, the I-V curve of leptin-induced outward currents is characterized by positive slope conductance and has an average reversal potential of -88 +/- 3 mV, suggesting that an increase in potassium conductance may underlie leptin-induced hyperpolarization. Most of the leptin-responsive DMNV neurons were identified as being parasympathetic preganglionic neurons. These results suggest that the DMNV is one of the central target sites of leptin, and leptin can regulate parasympathetic outflow from the DMNV by directly acting on the parasympathetic preganglionic neurons of the DMNV.     

Localization of cholinergic neurons in the cat lower brain stem

The localization of cholinergic neurons in the cat lower brain stem was determined immunocytochemically with a monoclonal antibody against choline acetyltransferase (ChAT), the acetylcholine synthesizing enzyme. ChAT-positive neurons were observed in four major cell groups: cranial nerve motor and special visceromotor neurons: parasympathetic preganglionic visceromotor neurons; neurons located in the ponto-mesencephalic tegmentum including area X (or pedunculopontine tegmental nucleus), nucleus laterodorsalis tegmenti (Ldt) of Castaldi, and peri-locus coeruleus alpha (peri-alpha); and neurons located in nucleus reticularis magnocellularis (Mc) and adjacent nucleus reticularis gigantocellularis (Gc) of the medulla.     

Evidence for a cell-specific action of Reelin in the spinal cord

Reelin, the extracellular matrix protein missing in reeler mice, plays an important role in neuronal migration in the central nervous system. We examined the migratory pathways of phenotypically identified spinal cord neurons to determine whether their positions were altered in reeler mutants. Interneurons and projection neurons containing choline acetyltransferase and/or NADPH diaphorase were studied in E12.5-E17.5 reeler and wild-type embryos, and their final locations were assessed postnatally. While three groups of dorsal horn interneurons migrated and differentiated normally in reeler mice, the migrations of both sympathetic (SPNs) and parasympathetic preganglionic neurons (PPNs) were aberrant in the mutants. Initially reeler and wild-type SPNs were detected laterally near somatic motor neurons, but by E13.5, many reeler SPNs had mismigrated medially. Postnatally, 79% of wild-type SPNs were found laterally, whereas in reeler, 92% of these neurons were positioned medially. At E13.5, both reeler and wild-type PPNs were found laterally, but by E14.5, reeler PPNs were scattered across the intermediate spinal cord while wild-type neurons correctly maintained their lateral location. By postnatal day 16, 97% of PPNs were positioned laterally in wild-type mice; in contrast, only 62% of PPNs were found laterally in mutant mice. In E12.5-E14.5 wild-type mice, Reelin-secreting cells were localized along the dorsal and medial borders of both groups of preganglionic neurons, but did not form a solid barrier. In contrast, Dab1, the intracellular adaptor protein thought to function in Reelin signaling, was expressed in cells having positions consistent with their identification as SPNs and PPNs. In combination, these findings suggest that, in the absence of Reelin, both groups of autonomic motor neurons migrate medially past their normal locations, while somatic motor neurons and cholinergic interneurons in thoracic and sacral segments are positioned normally. These results suggest that Reelin acts in a cell-specific manner on the migration of cholinergic spinal cord neurons.     

Musings on the wanderer: what's new in our understanding of vago-vagal reflex? IV. Current concepts of vagal efferent projections to the gut

Vagal efferents, consisting of distinct lower motor and preganglionic parasympathetic fibers, constitute the motor limb of vagally mediated reflexes. Arising from the nucleus ambiguus, vagal lower motor neurons (LMN) mediate reflexes involving striated muscles of the orad gut. LMNs provide cholinergic innervation to motor end plates that are inhibited by myenteric nitrergic neurons. Preganglionic neurons from the dorsal motor nucleus implement parasympathetic motor and secretory functions. Cholinergic preganglionic neurons form parallel inhibitory and excitatory vagal pathways to smooth muscle viscera and stimulate postganglionic neurons via nicotinic and muscarinic receptors. In turn, the postganglionic inhibitory neurons release ATP, VIP, and NO, whereas the excitatory neurons release ACh and substance P. Vagal motor effects are dependent on the viscera's intrinsic motor activity and the interaction between the inhibitory and excitatory vagal influences. These interactions help to explain the physiology of esophageal peristalsis, gastric motility, lower esophageal sphincter, and pyloric sphincter. Vagal secretory pathways are predominantly excitatory and involve ACh and VIP as the postganglionic excitatory neurotransmitters. Vagal effects on secretory functions are exerted either directly or via release of local mediators or circulating hormones.     

Chemical profile of vagal preganglionic motor cells innervating the airways in ferrets: the absence of noncholinergic neurons.

In ferrets, we investigated the presence of choline acetyltransferase (ChAT), vasoactive intestinal peptide (VIP), and markers for nitric oxide synthase (NOS) in preganglionic parasympathetic neurons innervating extrathoracic trachea and intrapulmonary airways. Cholera toxin beta-subunit, a retrograde axonal transganglionic tracer, was used to identify airway-related vagal preganglionic neurons. Double-labeling immunohistochemistry and confocal microscopy were employed to characterize the chemical nature of identified airway-related vagal preganglionic neurons at a single cell level. Physiological experiments were performed to determine whether activation of the VIP and ChAT coexpressing vagal preganglionic neurons plays a role in relaxation of precontracted airway smooth muscle tone after muscarinic receptor blockade. The results showed that 1) all identified vagal preganglionic neurons innervating extrathoracic and intrapulmonary airways are acetylcholine-producing cells, 2) cholinergic neurons innervating the airways coexpress ChAT and VIP but do not contain NOS, and 3) chemical stimulation of the rostral nucleus ambiguus had no significant effect on precontracted airway smooth muscle tone after muscarinic receptor blockade. These studies indicate that vagal preganglionic neurons are cholinergic in nature and coexpress VIP but do not contain NOS; their stimulation increases cholinergic outflow, without activation of inhibitory nonadrenergic, noncholinergic ganglionic neurons, stimulation of which induces airway smooth muscle relaxation. Furthermore, these studies do not support the possibility of direct inhibitory innervation of airway smooth muscle by vagal preganglionic fibers that contain VIP.     

Adrenergic innervation of the dorsal vagal motor nucleus: possible involvement in inhibitory control of gastric acid and pancreatic insulin secretion

Summary Morphological and physiological approaches were used to investigate the possible role of an adrenergic innervation of the dorsal vagal complex in the control of basal gastric acid and pancreatic insulin secretion in the rat. The use of retrograde-tracing methods with injections of True Blue or of wheat-germ agglutinin into the stomach or pancreas first confirmed that most vagal preganglionic neurons innervating these two viscera are localized in the dorsal motor nucleus of the vagus, a number of them connected to both viscera. Light- and electron-microscopic investigation of the organization of adrenergic neuronal structures immunoreactive to phenylethanolamine-N-methyltransferase within this medullary nucleus further revealed: (i) that adrenergic axons establish profuse synaptic connections of the symmetrical type with perikarya and dendrites of this nucleus, and (ii) that several of these adrenergic fibers are connected with retrogradely labeled neurons innervating the stomach and/or pancreas. Lastly, measurements of basal gastric acid output and plasma insulin clearly indicated that both visceral secretions are rapidly and conspicuously decreased by local infusion of 2 nM adrenaline within the dorsal vagal complex. Taken together, these data strongly suggest that the adrenergic innervation of the dorsal medulla oblongata is involved in direct synaptic inhibition of the parasympathetic preganglionic neurons of the vagus that control secretion of gastric acid and pancreatic insulin.     

Axons of sacral preganglionic neurons in the cat: II. Axon collaterals

Axon collaterals were identified in 21 of 24 preganglionic neurons in the lateral band of the sacral parasympathetic nucleus of the cat. Following the intracellular injection of HRP or neurobiotin the axons from 20 of these neurons were followed and 53 primary axon collaterals were found to originate from unmyelinated segments and from nodes of Ranvier. Detailed mapping done in the five best labeled cells showed bilateral axon collaterals distributions up to 25,000 μm in length with 950 varicosities and unilateral distributions up to 12,561 μm with 491 varicosities. The axon collaterals appeared to be unmyelinated, which was confirmed at EM, and were small in diameter (average 0.3 μm). Varicosities were located mostly in laminae I, V, VII, VIII and X and in the lateral funiculi. Most varicosities were not in contact with visible structures but some were seen in close apposition to Nissl stained somata and proximal dendrites. Varicosities had average minor diameters of 1.3 μm and major diameters of 2.3 μm. Most were boutons en passant while 10–20% were boutons termineaux. EM revealed axodendritic and axoaxonic synapses formed by varicosities and by the axons between varicosities. It is estimated that the most extensive of these axon collaterals systems may contact over 200 spinal neurons in multiple locations. These data lead to the conclusion that sacral preganglionic neurons have multiple functions within the spinal cord in addition to serving their target organ. As most preganglionic neurons in this location innervate the urinary bladder, it is possible that bladder preganglionic neurons have multiple functions.     

Transneuronal labeling of neurons in the adult rat central nervous system following inoculation of pseudorabies virus into the colon

Transneuronal tracing with pseudorabies virus (PRV) was used to identify sites in the central nervous system involved in the neural control of colon function. PRV-immunoreactive (IR) cells were primarily localized to the caudal lumbosacral (L6-S1) and caudal thoracic-rostral lumbar (T13-L1) spinal segments with the distribution varying according to survival time (72-96 h). In the lumbosacral spinal cord at all time points examined, significantly (PА.005) greater numbers of PRV-IR cells were present in the region of the sacral parasympathetic nucleus (SPN) of the S1 spinal segment compared to that of the L6 segment. These studies also revealed morphologically distinct cell types with a differential distribution (probably interneurons and preganglionic parasympathetic neurons) in the region of the SPN in the L6-S1 spinal segments following colon inoculation. PRV-labeled neurons were located at various levels of the neuraxis and at many sites had a distribution similar to that following injection of virus to other urogenital organs. However, some unique sites in the dorsal motor nucleus of the vagus, nucleus of the solitary tract, nucleus ambiguus and area postrema were also identified. To determine if labeling in these caudal medullary sites was mediated by spinal or vagal pathways, the colon was inoculated with PRV in animals with a complete spinal cord (T8) transection (5-7 days prior). Following spinal transection, PRV-infected cells were detected in the same caudal medullary regions; however, labeling in other regions (e.g., Barrington's nucleus) was eliminated or significantly reduced. These studies have yielded several novel observations concerning the central neural control of colonic function: (1) the preganglionic efferent and primary afferent innervation of the colon arises primarily from the S1 spinal segment; (2) the distribution of PRV-infected neurons in the central nervous system following colon inoculation was similar to that following PRV inoculation of other urogenital organs; (3) Barrington's nucleus, which has been identified previously as the pontine micturition center, may have a role in colonic function; and (4) PRV infection in Barrington's nucleus following colon inoculation is mediated by bulbospinal pathways whereas labeling in caudal medullary regions is mediated, at least in part, by vagal pathways.     

A Novel Central Pathway Links Arterial Baroreceptors and Pontine Parasympathetic Neurons in Cerebrovascular Control

1. We tested the hypothesis that arterial baroreceptor reflexes modulate cerebrovascular tone through a pathway that connects the cardiovascular nucleus tractus solitarii with parasympathetic preganglionic neurons in the pons.2. Anesthetized rats were used in all studies. Laser flowmetry was used to measure cerebral blood flow. We assessed cerebrovascular responses to increases in arterial blood pressure in animals with lesions of baroreceptor nerves, the nucleus tractus solitarii itself, the pontine preganglionic parasympathetic neurons, or the parasympathetic ganglionic nerves to the cerebral vessels. Similar assessments were made in animals after blockade of synthesis of nitric oxide, which is released by the parasympathetic nerves from the pterygopalatine ganglia. Finally the effects on cerebral blood flow of glutamate stimulation of pontine preganglionic parasympathetic neurons were evaluated.3. We found that lesions at any one of the sites in the putative pathway or interruption of nitric oxide synthesis led to prolongation of autoregulation as mean arterial pressure was increased to levels as high as 200 mmHg. Conversely, stimulation of pontine parasympathetic preganglionic neurons led to cerebral vasodilatation. The second series of studies utilized classic anatomical tracing methods to determine at the light and electron microscopic level whether neurons in the cardiovascular nucleus tractus solitarii, the site of termination of baroreceptor afferents, projected to the pontine preganglionic neurons. Fibers were traced with anterograde tracer from the nucleus tractus solitarii to the pons and with retrograde tracer from the pons to the nucleus tractus solitarii. Using double labeling techniques we further studied synapses made between labeled projections from the nucleus tractus solitarii and preganglionic neurons that were themselves labeled with retrograde tracer placed into the pterygopalatine ganglion.4. These anatomical studies showed that the nucleus tractus solitarii directly projects to pontine preganglionic neurons and makes asymmetric, seemingly excitatory, synapses with those neurons. These studies provide strong evidence that arterial baroreceptors may modulate cerebral blood flow through direct connections with pontine parasympathetic neurons. Further study is needed to clarify the role this pathway plays in integrative physiology.     

Nesfatin-1 inhibits gastric acid secretion via a central vagal mechanism in rats

Nesfatin-1, a novel hypothalamic peptide, inhibits nocturnal feeding behavior and gastrointestinal motility in rodents. The effects of nesfatin-1 on gastrointestinal secretory function, including gastric acid production, have not been evaluated. Nesfatin-1 was injected into the fourth intracerebral ventricle (4V) of chronically cannulated rats to identify a nesfatin dose sufficient to inhibit food intake. Nesfatin-1 (2 μg) inhibited dark-phase food intake, in a dose-dependent fashion, for >3 h. Gastric acid production was evaluated in urethane-anesthetized rats. Nesfatin-1 (2 μg) was introduced via the 4V following endocrine stimulation of gastric acid secretion by pentagastrin (2 μg·kg(-1)·h(-1) iv), vagal stimulation with 2-deoxy-d-glucose (200 mg/kg sc), or no stimulus. Gastric secretions were collected via gastric cannula and neutralized by titration to determine acid content. Nesfatin-1 did not affect basal and pentagastrin-stimulated gastric acid secretion, whereas 2-deoxy-d-glucose-stimulated gastric acid production was inhibited by nesfatin-1 in a dose-dependent manner. c-Fos immunofluorescence in brain sections was used to evaluate in vivo neuronal activation by nesfatin-1 administered via the 4V. Nesfatin-1 caused activation of efferent vagal neurons, as evidenced by a 16-fold increase in the mean number of c-Fos-positive neurons in the dorsal motor nucleus of the vagus (DMNV) in nesfatin-1-treated animals vs. controls (P < 0.01). Finally, nesfatin-induced Ca(2+) signaling was evaluated in primary cultured DMNV neurons from neonatal rats. Nesfatin-1 caused dose-dependent Ca(2+) increments in 95% of cultured DMNV neurons. These studies demonstrate that central administration of nesfatin-1, at doses sufficient to inhibit food intake, results in inhibition of vagally stimulated secretion of gastric acid. Nesfatin-1 activates DMNV efferent vagal neurons in vivo and triggers Ca(2+) signaling in cultured DMNV neurons.     

GRP mediates an inhibitory response of gut-related vagal motor neurons to PVN stimulation

We previously characterized neurons in the dorsal motor nucleus of the vagus (DMNV) that were modulated by electrical stimulation of the PVN and by gastrointestinal distention. Bombesin has been identified in a subset of PVN neurons projecting to the DMNV. It is currently unknown whether this neurotransmitter is involved in descending communication from PVN to DMNV neurons. In this study we determined whether the specific bombesin antagonist, N-acetyl-GRP(20-26), influenced (1) the basal firing rate of DMNV neurons and (2) the response to electrical current stimulation of the PVN. Our results indicate that N-acetyl-GRP(20-26), significantly attenuated the inhibitory response of DMNV neurons to PVN stimulation. These results provide a possible mechanism by which bombesin regulates gastrointestinal function, body temperature homeostasis, and feeding behaviors.     

Spatial organization of neurons in the dorsal motor nucleus of the vagus synapsing with intragastric cholinergic and nitric oxide/VIP neurons in the rat

The dorsal motor nucleus of the vagus (DMV) contains preganglionic neurons that control gastric motility and secretion. Stimulation of different parts of the DMV results in a decrease or an increase in gastric motor activities, suggesting a spatial organization of vagal preganglionic neurons in the DMV. Little is known about how these preganglionic neurons in the DMV synapse with different groups of intragastric motor neurons to mediate contraction or relaxation of the stomach. We used pharmacological and immunohistochemical methods to characterize intragastric neural pathways involved in mediating gastric contraction and relaxation in rats. Microinjections of L-glutamate (L-Glu) into the rostral or caudal DMV produced gastric contraction and relaxation, respectively, in a dose-related manner. Intravenous infusion of hexamethonium blocked these actions, suggesting mediation via preganglionic cholinergic pathways. Atropine inhibited gastric contraction by 85.5 +/- 4.5%. Gastric relaxation was reduced by intravenous administration of N(G)-nitro-L-arginine methyl ester (L-NAME; 52.5 +/- 11.9%) or VIP antagonist (56.3 +/- 14.9%). Combined administration of L-NAME and VIP antagonist inhibited gastric relaxation evoked by L-Glu (87.8 +/- 4.3%). Immunohistochemical studies demonstrated choline acetyltransferase immunoreactivity in response to L-Glu microinjection into the rostral DMV in 88% of c-Fos-positive intragastric myenteric neurons. Microinjection of L-Glu into the caudal DMV evoked expression of nitric oxide (NO) synthase and VIP immunoreactivity in 81 and 39%, respectively, of all c-Fos-positive intragastric myenteric neurons. These data indicate spatial organization of the DMV. Depending on the location, microinjection of L-Glu into the DMV may stimulate intragastric myenteric cholinergic neurons or NO/VIP neurons to mediate gastric contraction and relaxation.     

Differential development of cholinergic neurons from cranial and trunk neural crest cells in vitro

Several studies have suggested that the development of cholinergic properties in cranial parasympathetic neurons is determined by these cells' axial level of origin in the neural crest. All cranial parasympathetic neurons normally derive from cranial neural crest. Trunk neural crest cells give rise to sympathetic neurons, most of which are noradrenergic. To determine if there is an intrinsic difference in the ability of cranial and trunk neural crest cells to form cholinergic neurons, we have compared the development of choline acetyltransferase (ChAT)-immunoreactive cells in explants of quail cranial and trunk neural crest in vitro. Both cranial and trunk neural crest explants gave rise to ChAT-immunoreactive cells in vitro. In both types of cultures, some of the ChAT-positive cells also expressed immunoreactivity for the catecholamine synthetic enzyme tyrosine hydroxylase. However, several differences were seen between cranial and trunk cultures. First, ChAT-immunoreactive cells appeared two days earlier in cranial than in trunk cultures. Second, cranial cultures contained a higher proportion of ChAT-immunoreactive cells. Finally, a subpopulation of the ChAT-immunoreactive cells in cranial cultures exhibited neuronal traits, including neurofilament immunoreactivity. In contrast, neurofilament-immunoreactive cells were not seen in trunk cultures. These results suggest that premigratory cranial and trunk neural crest cells differ in their ability to form cholinergic neurons.     

Axons of sacral preganglionic neurons in the cat: I. Origin,initial segment,and myelination

Parasympathetic preganglionic neurons in the cat sacral spinal cord innervate intraspinal neurons and pelvic target organs. Retrograde tracing studies have revealed little of the morphology of their axons including their origin, initial segments, or their myelin, due to methodological limitations. Intracellular labeling of single neurons with neurobiotin or HRP has overcome these problems. Axons were studied in 24 preganglionic neurons. In 21 neurons the axon originated as a branch of a dendrite, without a detectable axon hillock, at distances from the soma ranging from 10 to 110 μm (average 34.1 μm ). In 3 neurons the axon was derived from the soma. Initial segments, present in all cells, ranged from 15 to 40 μm (average 26.8 μm). Nearly all axons followed the initial segment with unmyelinated segments that varied between 59 to 630 μm, followed by myelin and nodes of Ranvier. Internodal distances were variable and relatively short (average 93 μm). Axonal diameters measured over the intraspinal course in 18 axons averaged 1.3 μm (range 0.6–2.4 μm) and were relatively constant compared with other neurons. Spine-like protrusions were observed on the initial segments of 12 cells. Axon collaterals originated from unmyelinated sections and nodes of Ranvier. Antidromic action potentials showing initial segment, soma-dendritic inflections, did not differentiate between soma-derived and dendrite-derived axons. The data suggest that axons originating from a dendrite are the normal structure of preganglionic neurons in the lateral sacral parasympathetic nucleus. It is proposed that the particular structure of these axons may be part of a timing mechanism that coordinates preganglionic neurons with other spinal neurons involved in target organ reflexes.     

Immunohistochemical characterisation of sympathetic and parasympathetic pelvic neurons projecting to the distal colon in the male rat

The pelvic ganglia are mixed ganglia containing both sympathetic and parasympathetic neurons that receive spinal input via the hypogastric (lumbar cord) and pelvic nerves (sacral cord), respectively. A recent study has utilised immunohistochemistry against synaptophysin (a protein associated with small vesicles) to visualise the preganglionic terminals in these ganglia. By selectively cutting the hypogastric or pelvic nerves and allowing subsequent terminal degeneration, the populations of parasympathetic and sympathetic preganglionic terminals, respectively, can be visualised. The present study has used this method in conjunction with retrograde labelling of pelvic neurons from the distal colon and double label immunofluorescence against tyrosine hydroxylase and vasoactive intestinal polypeptide (VIP) to identify and characterise the sympathetic and parasympathetic neurons projecting to the distal colon from the major pelvic ganglia of the male rat. Approximately equal numbers of distal colonic-projecting pelvic neurons are sympathetic and parasympathetic. Almost all noradrenergic neurons are sympathetic. Of the VIP neurons that project to the distal colon approximately one third are sympathetic, one third parasympathetic and the remaining third are possibly innervated by both the lumbar and sacral cord. Extrapolation from our results also suggests that the majority of non-noradrenergic neuropeptide Y neurons (which are known to comprise the remainder of the neurons) are parasympathetic. These studies have demonstrated that the pelvic ganglia are a major source of sympathetic innervation to the distal bowel and have further shown that the distal colon is another target for the non-noradrenergic sympathetic neurons of the pelvic ganglia.     

The organization of cholinergic neurons in the mesencephalon of the eel,Anguilla anguilla,as determined by choline acetyltransferase immunohistochemistry and acetylcholinesterase enzyme histochemistry

Cholinergic systems in the midbrain of the eel were identified by using histochemical procedures for the demonstration of the enzymes choline acetyltransferase (ChAT) and acetylcholinesterase. Neurons detected by both methods are located in the stratum periventriculare of the tectum, cranial motor nuclei III and IV, nucleus isthmi, nucleus gustatorius secundarius, nucleus reticularis superior, and nucleus lateralis valvulae. Some projections of these cell groups were studied by injecting horseradish peroxidase into selected brain regions. Cholinergic neurons make up about 10% of the neurons in the stratum periventriculare of the tectum and are a subset of the type-XIV neurons. Neurons in n. isthmi project primarily to the ipsilateral tectum; some cholinergic isthmal neurons project to n. pretectalis superficialis. A few ChAT-positive axons, perhaps belonging to the tectopetal system, were observed in the optic nerve. The cholinergic neurons of n. gustatorius secundarious project to the inferior lobes of the hypothalamus. The neurons of the superior reticular nucleus are a cholinergic subset of the superior reticular formation. Their axons project rostrally, probably to the thalamus and pretectum. The findings are discussed in relation to functional features of the mesencephalon, particularly in relation to locomotory control.     

Possible mechanisms of the anterior limbic cortex influence on the neuron activity of the vago-solitary complex

In ananesthetized cats, neurons of the nucleus of the tractus solitarius (NTS) and the dorsal motor nucleus of the vagus nerve (DMNV) revealed phasic excitatory responses to separate single vagal and cortical stimuli. Stimulation of the anterior limbic cortex combined with vagal stimulation resulted in inhibitory or excitatory modification of the vagal induced responses of the NTS and DMNV neurons. The data obtained suggest that complete inhibitory effects are related to general cortical mechanisms of control of the functional state of the brain stem visceral neurons. Selective inhibition of the vagal induced responses by limbic cortex stimulation is due to particular cortical mechanisms of the visceral sensory transmission control via the NTS neurons.     

Coexistence of neurons integrating urinary bladder activity and pelvic nerve activity in the same cardiovascular areas of the pontomedulla in cats

The present study examines the coexistence of neurons in the same cardiovascular point of the pontomedulla that integrates urinary bladder (UB) motility, and pelvic nerve activity (PNA). Microinjection of monosodium L-glutamate (Glu) into the locus coeruleus (LC), the gigantocellular tegmental field (FTG), the rostral ventrolateral medulla (RVLM), and the dorsomedial medulla (DM) produced pressor responses, whereas injection into the lateral tegmental field (FTL), the nucleus of tractus solitarii (NTS), and the caudal ventrolateral medulla (CVLM) produced depressor responses. However, microinjection of Glu into the dorsomotor nucleus of the vagus (DMV) and the ambiguus nucleus (AN), where the vagus nerve originates, produced marked bradycardia. Many of these cardiovascular responses were accompanied by increased, or decreased parasympathetic PNA. In six animals, sympathetic renal nerve activity (RNA) and PNA also increased simultaneously during the pressor response. The present study also examines the connection between the DMV-AN and the sacral intermediolateral column (IML), where parasympathetic preganglionic neurons (PGNs) of the pelvic nerve located. Biotinylated dextran amine (BDA), an anterograde tracer, was iontophoretically injected into the DMV or AN. No labelled terminal or neuron was detected in the sacral IML, but labelled terminals were observed in the bilateral LC, and also in the bilateral sides of the FTG, FTL, RVLM, DM, and CVLM. These results suggest that neurons of the DMV and/or AN may indirectly regulate the sacral parasympathetic PGNs through the LC for supraspinal control of the pelvic nerve. Furthermore, these results also suggest the coexistence of multiple autonomic integrating mechanisms of different kinds within various cardiovascular areas of the pontomedulla.     

Effect of preganglionic sympathectomy on the cholinesterase activity in the superior cervical ganglia of rats and hamsters

Summary By employing biochemical assay and histochemical enzyme techniques the effect of preganglionic sympathectomy on the cholinesterase (ChE) activity in the superior cervical ganglia of rats and hamsters was investigated. Biochemical assays indicate that the ChE activity in the superior cervical ganglia of adult rats and hamsters is 57.19 and 28.63 respectively (expressed in u moles acetylcholine hydrolyzed per min per g of tissue); two weeks after preganglionic denervation, about 50% and 60% of ChE activity are lost respectively. Histochemical enzyme examination reveals that in the rat superior cervical ganglion, the majority of the neurons are adrenergic with weak to moderate acetylcholinesterase (AChE) reaction and the minority of the neurons are cholinergic with strong AChE activity, while only one type of adrenergic neurons exhibits a weak AChE activity in the hamster superior cervical ganglion. The AChE activity is localized in the perinuclear area, in the cisternae of the rough surfaced endoplasmic reticulum, in the Golgi complex and on the plasma membrane of the hamster's neurons; it is mainly localized in the cisternae of the rough surfaced endoplasmic reticulum of the rat's neurons. AChE reaction product is also detected on the axolemmal membranes of the preganglionic nerve fibers in the sympathetic ganglia of rats and hamsters.After preganglionic sympathectomy, the AChE activity in the adrenergic neurons and in the preganglionic unmyelinated nerve fibers is markedly reduced, whereas the cholinergic neurons and preganglionic myelinated nerve fibers remain unchanged. On the basis of these results two conclusions have been reached: (1) The fact that strong AChE activity localized in the cholinergic neurons and preganglionic myelinated fibers is not influenced by denervation, suggests that these structures are able to produce AChE. (2) The reduction of AChE activity in the rat and hamster superior cervical ganglia two weeks after preganglionic denervation, observed by histochemical examination, can be correlated with a concomitant measurable reduction determined by biochemical assays.Supported in part by a grant from the National Science Council, Republic of China. The author wishes to express his gratitude to the Department of Pharmacology, College of Medicine, National Taiwan University, for the use of its equipment for biochemical assays